Project Grant R01AI179743

THE EPIGENETICS OF CYTOPLASMIC INCOMPATIBILITY - PROJECT SUMMARY: WOLBACHIA ARE A GENUS OF ENDOSYMBIOTIC BACTERIA THAT COMPRISE A PROMISING, COST-EFFECTIVE TOOL TO CURB ARBOVIRAL TRANSMISSION BASED ON TWO KEY FACETS. FIRST, WOLBACHIA BLOCK PATHOGENIC RNA VIRUSES BY INHIBITING THEIR REPLICATION IN ARTHROPODS. SECOND, WOLBACHIA SELFISHLY ALTER SPERM AND EGG VIA A PROCESS TERMED CYTOPLASMIC INCOMPATIBILITY (CI) THAT CAN DRIVE THE BACTERIA INTO HOST POPULATIONS. CI IS EXPRESSED AS EMBRYONIC LETHALITY IN CROSSES BETWEEN INFECTED MALES AND UNINFECTED FEMALES, BUT THIS LETHALITY IS RESCUED IN CROSSES BETWEEN INFECTED MALES AND INFECTED FEMALES, WHICH ARE THE TRANSMITTING SEX OF WOLBACHIA. CONSEQUENTLY, CI IS DEPLOYED IN FIELD TRIALS TO EITHER SUPPRESS MOSQUITO POPULATION SIZES OR REPLACE UNINFECTED POPULATIONS WITH INFECTED INDIVIDUALS RESISTANT TO ARBOVIRAL INFECTION. WE RECENTLY DISCOVERED PRE-FERTILIZATION IMPAIRMENTS TO SPERM GENOME INTEGRITY THAT UNDERPIN THE CI DRIVE. THE CI-CAUSING CIF PROTEINS CIFA AND CIFB FROM A PROPHAGE REGION IN WMEL INTERACT WITH DEVELOPING SPERMATID NUCLEI TO ALTER THE EPIGENETICALLY-CONTROLLED, HISTONE-TO-PROTAMINE TRANSITION, NAMELY BY INCREASING HISTONE RETENTION IN DEVELOPING SPERMATIDS AND DECREASING PROTAMINE LEVELS IN THE MATURE SPERM. THE CIF PROTEINS DO NOT PATERNALLY TRANSFER TO THE FERTILIZED EMBRYOS, INSTEAD THE MODIFIED SPERM IN TESTES WITH IMPAIRED CHROMATIN INTEGRITY TRANSFERS TO BESTOW CI. DESPITE DECADES OF INTENSE RESEARCH AND CURRENT APPLICATIONS TO VECTOR CONTROL, THE MECHANISTIC DETAILS OF THESE PRE-FERTILIZATION IMPAIRMENTS REMAIN A CENTRAL ENIGMA AND ARE ONLY NOW SUBJECT TO INVESTIGATION. HERE, WE PROPOSE THE FIRST, IN-DEPTH EXAMINATION OF CI-DEFINING HISTONE MARKS AND PROTAMINE TYPES TO TEST THE CENTRAL HYPOTHESIS THAT THE CIF PROTEINS ALTER EPIGENETIC EVENTS DURING GAMETOGENESIS TO CAUSE CI AND RESCUE. IN AIM 1, WE WILL USE CUT & TAG CHROMATIN PROFILING TO DETERMINE CI- DEFINING, PRE-FERTILIZATION CHANGES IN SPERM HISTONE MODIFICATIONS CAUSING HISTONE RETENTION IN WMEL-INFECTED DROSOPHILA MELANOGASTER AND AEDES AEGYPTI. WE WILL ALSO TEST THE ESSENTIALITY OF THESE EPIGENETIC CHANGES TO CI BY CHEMICALLY INHIBITING HISTONE MODIFIER ENZYMES. IN AIM 2, WE WILL INVESTIGATE THE TYPE AND ABUNDANCE OF CI- DEFINING PROTAMINES THAT ARE DEPLETED IN MATURE SPERM. KNOCKOUT AND KNOCKDOWN TRANSGENIC EXPERIMENTS WILL LINK CI-DEFINING EMBRYONIC DEATH WITH THE NECESSITY OF SPECIFIC SPERM PROTAMINE GENES. FINALLY IN AIM 3, WE WILL EVALUATE RESCUE-DEFINING CHANGES IN THE GAMETIC CHROMATIN BY FIRST CHARACTERIZING MATERNAL CHANGES IN HISTONE TRANSCRIPTS AND PROTEIN ABUNDANCE DURING OOGENESIS. WE WILL THEN DETERMINE POST-FERTILIZATION EPIGENETIC CHANGES IN SPERM DNA THAT ORCHESTRATE PATERNAL CHROMATIN DECONDENSATION FOR EMBRYONIC VIABILITY. WE WILL ALSO VALIDATE THE ESSENTIALITY OF EPIGENETIC MODIFIERS TO RESCUE BY INHIBITING THEIR ENZYMATIC ACTIVITY. DESPITE THE RISING INTEREST IN DEPLOYING WOLBACHIA TO CURB ARBOVIRUS TRANSMISSION, STUDIES HAVE NOT YET YIELDED A REFINED, MOLECULAR BREAKTHROUGH UNCOVERING THE HOST EPIGENETICS DRIVING WOLBACHIA. IF SUCCESSFUL, THE PROPOSED RESEARCH WILL (I) REVEAL THE INCIPIENT, HOST EPIGENETIC EVENTS UNDERLYING THE CI DRIVE SYSTEM AT THE HEART OF VECTOR CONTROL STRATEGIES AND (II) PROVIDE A SIGNIFICANT GATEWAY TO EVENTUALLY ENGINEER CI AND RESCUE VIA THE HOST ARTHROPOD ITSELF.